4D Lattice Light-Sheet Organoid Imaging
I use endogenously GFP/RFP tagged stem cells to grow tissues (organoids) and then image them live with the lattice light-sheet microscope (LLSM). Shown below is a young brain organoid and a polarized intestinal organoid. LLSM imaging is done at superresolution with seconds frame rates. The resulting large datasets (terabytes) are analyzed using my custom analysis software pyLattice.
pyLattice - Lattice Light-Sheet Big Data Analytics
pyLattice is a python library for advanced lattice light-sheet image analysis. Collaborate with us on gitHub and read about how we solve big data analytics challenges when we analyze big LLSM datasets.
Scope Engineering and Optical Tweezing
I combined confocal microscopy with optical trapping to measure force and fluorescence of proteins acting on small membrane nanotubes (pulled from e.g. a giant unilamellar vesicles (GUV)). These nanotubes allow studying protein-membrane biophysics in tubular, highly curved topologies. They resemble vesicle necks found e.g. in normal topology budding (e.g. Clathrin mediated endocytosis) and reverse topology budding (ESCRT system).
The Confleezers: Left:The Confleezers 1.0 that combines confocal microscopy with optical tweezing. Right: Membrane nanotubes pulled from model membranes using micromanipulators (top) and the trap of the Confleezers (bottom).
See our applications on the ESCRT system and our review:
J Schöneberg, S Yan, M Righini, M R Pavlin, I-H Lee, L-A Carlson, A H Bahrami, D H Goldman, X Ren, G Hummer, C Bustamante, J Hurley. (2018) ATP-dependent force generation and membrane scission by ESCRT-III and Vps4. bioRxiv, doi: https://doi.org/10.1101/262170.
J Schöneberg*, Lee IH*, Iwasa JH, Hurley JH, (2016) Reverse-topology membrane scission by the ESCRT proteins. Nature Reviews Molecular Cell Biology, doi: 10.1038/nrm.2016.12. (*equal contribution).
J Schöneberg*, M Lehmann*, A Ullrich, Y Posor, W-T Lo, G Lichtner, J Schmoranzer, V Haucke, F Noé, (2017) Lipid-mediated PX-BAR domain recruitment couples local membrane constriction to endocytic vesicle fission. Nature Communications 8, 15873 doi: 10.1038/ncomms15873. (*equal contribution).
See our contributions to the rod cell visual cascade: Structure 2015: Higher-order architecture of rhodopsin in intact photoreceptors and its implication for phototransduction kinetics. Biophysical Journal 2014: Explicit Spatiotemporal Simulation of Receptor-G Protein Coupling in Rod Cell Disk Membranes.
ReaDDy is a particle-based reaction-diffusion simulation software. It features
- simulation in and on arbitrary geometries (i.e. 3D, 2D, spherical,...),
- spatial confinement (walls, boxes, tubes, ...),
- excluded volume of particles (crowding effects) and
- particle-particle interaction potentials (repulsion, attraction, clustering, ...).